DNA replication and DNA repair in Acholeplasma laidlawii would be studied with the following specific aims: The initiation process of DNA replication would be studied using amino acid starvation and antibiotic inhibition of reinitiation. Isolation of restriction fragments containing the origin would be done using restriction fragment analysis of cells radioactively labeled at the origin. Such fragments would be cloned in E. coli K12 strains, thereby creating origin plasmids for detailed structural studies of origin DNA and for in vitro studies of initiation. Studies of the elongation process would center on isolation and characterization of Okazaki fragments, on the rate of chain elongation, and on autoradiographic evidence for bidirectional vs. unidirectional replication. The properties of A. laidlawii folded chromosomes as related to the DNA replication cycle would be analyzed via sedimentation and electron microscopic techniques. A. laidlawii plasmids would be characterized, and enzymes related to DNA structure and function would be sought and isolated. Genes for such enzymes would be isolated via cloning of A. laidlawii DNA restriction fragments in E. coli K12 strains auxotrophic for DNA functions.